In vitro studies of the role of P2X4 receptors on inflammatory processes involved in Parkinsons Disease

• Grant number: 22/01304-2
• Support Opportunities: Scholarships in Brazil - Master
• Start date: September 01, 2022
• End date: March 02, 2024
• Field of knowledge: Biological Sciences - Pharmacology - Biochemical and Molecular Pharmacology
• Principal Investigator: Alexander Henning Ulrich
• Grantee: Beatriz de Andrade de Faria
• Host Institution: Instituto de Química (IQ). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated research grant: 18/07366-4 - Purine and kinin receptors as targets of study and therapeutic interventions in neurological diseases, AP.TEM

Abstract

Parkinson's disease is a neurodegenerative disease that involves the death of dopaminergic neurons of the nigrostriatal pathway and the formation of protein agglomerates composed by ±-sinuclein (Lewy bodies). Neurodegeneration is strongly linked to neuroinflammation, mainly caused by microglial cells of the nigrostriatal pathway. Therefore, the project proposes the in vitro study of a neuroinflammatory pathway in microglia mediated by P2X4 purinergic receptors (P2X4R). For that, the human SH-SY5Y cell line will be differentiated into dopaminergic neurons and exposed to 6-OHDA for a Parkinson's disease in vitro model. Following this step, microglial human C20 cells will be exposed to the medium of 6-OHDA-exposed SH-SY5Y cells, rich in ATP. Then, analysis of microglial activation and activation of microglial P2X4R by ATP will be performed. For this purpose, C20 cell cultures will be exposed to drugs such as 5-BDBD, a P2X4R antagonist, and ivermectin, a P2X4R positive allosteric modulator. The pathway supposedly intermediated by the P2X4R involves an increase in intracellular concentrations of calcium ions (Ca2+) that have access to the cell through an ion permeable channel in activated P2X4 receptors. These ions cause damage to the mitochondria that then starts producing reactive oxygen species capable of activating the NLRP3 inflammasome. Once activated, the inflammasome activates cytokine molecules, by cleaving pro-interleukin-18 and pro-interleukin-1², into their final and pro-inflammatory forms, interleukin-18 and interleukin-1².